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An introduction to virology

introduction to viruses
structure and classification
basic virology
clinical virology

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An introduction to virology

  1. 1. 1
  2. 2. VIROLOGY (The study of Viruses) 2 Dr. Kaveh Haratian Department of Microbiology and Immunology Alborz University of Medical Sciences Fall 1394
  3. 3. 3 OUTLINE • introduction to viruses structure and classification • basic virology • clinical virology
  4. 4. In VIROLOGY we notice:  different structure  different method of replication  implications for  diagnosis  treatment  prevention 4
  5. 5. CONTROL METHODS  INVOLVE KNOWLEDGE OF:  RESERVOIRS  MODE OF TRANSMISSION  METHODS TO INACTIVATE VIRUS OF INTEREST  VACCINES  ANTI-VIRAL DRUGS  DEVELOPMENT OF DRUG RESISTANCE 5
  6. 6. EMERGING VIRAL DISEASES  Some new global examples:  HIV/AIDS  Hantavirus pulmonary syndrome (HPS)  West Nile encephalitis (WNV)  Severe acute respiratory syndrome (SARS)  Monkey pox  Human metapneumovirus  Ebola hemorrhagic disease 6
  7. 7. Consequences of viral infections  50% of all absenteeism  Children:  7 or more viral infections per year that involve a visit to a physician 7
  8. 8. Consequences of viral infections  Suffering, followed by recovery  Persistent disease  Fatal disease  Congenital disease  Contributory factor in cancer  Contributory factor in other diseases 8
  9. 9. SOME ARE ASYMPTOMATIC! 9
  10. 10. VIRUSES CAN BE USEFUL  VACCINE DEVELOPMENT  GENE THERAPY  TOOLS TO INVESTIGATE HOST CELLS 10
  11. 11. WHAT ARE VIRUSES? 11 “A PIECE OF BAD NEWS WRAPPED UP IN A PROTEIN”
  12. 12. WHAT ARE VIRUSES?  NUCLEIC ACID GENOME:  DNA OR RNA  PROTEIN COAT  PROTECTION, ENTRY  LIPID ENVELOPE IN SOME VIRUSES  SMALL  (20-400nm)  OBLIGATE INTRACELLULAR PARASITES 12
  13. 13. 13 Virus particle = virion White, DO and Fenner, FJ. Medical Virology, 4th Ed. 1994
  14. 14. 14Koneman et al. Color Atlas and Textbook of Microbiology 5th Ed. 1997
  15. 15. Virus versus Virion  Virus is a broad general term for any aspect of the infectious agent and includes: • the infectious or inactivated virus particle • viral nucleic acid and protein in the infected cell  Virion is the physical particle in the extra- cellular phase which is able to spread to new host cells; complete intact virus particle
  16. 16. 16 Growth on artificial media Division by binary fission Contain DNA and RNA Contain protein synthesi s machine ry Contain muramic acid Sensitiv e to antibioti cs Bacteria often yes yes yes often yes Viruses never no Either DNA or RNA no* no no * The arenavirus family appears to ‘accidentally’ package ribosomes, but these appear to play no role in protein synthesis.
  17. 17. CONSEQUENCES  NO BROAD RANGE ANTIBIOTICS  HEAVILY PARASITIC ON HOST CELL  NEED TO LOOK FOR WEAK LINK 17
  18. 18. HOST RANGE MAY BE WIDE OR NARROW MAY BE INSECT/ANIMAL, INSECT/PLANT DO NOT CROSS EUCARYOTE / PROCARYOTE BOUNDARY 18
  19. 19. 19 FACTORS AFFECTING HOST RANGE - CELL SURFACE RECEPTORS
  20. 20. FACTORS AFFECTING HOST RANGE  CELL SURFACE RECEPTORS 20 • AVAILABILITY OF REPLICATION MACHINERY • ABILITY TO GET OUT OF CELL AND SPREAD • HOST ANTI-VIRAL RESPONSE
  21. 21. VIRAL STRUCTURE : SOME TERMINOLOGY  virus particle = virion  protein which coats the genome = capsid  capsid usually symmetrical  capsid + genome = nucleocapsid  may have an envelope 21
  22. 22. ICOSAHEDRAL SYMMETRY  20 faces  12 vertices 22 http://www.tulane.edu/~dmsander/WWW/Video/Video.html
  23. 23. ICOSAHEDRAL SYMMETRY 23
  24. 24. ICOSAHEDRAL SYMMETRY 24
  25. 25. ICOSAHEDRAL SYMMETRY 25
  26. 26. ICOSAHEDRAL SYMMETRY 26
  27. 27. 27
  28. 28. 28
  29. 29. ICOSAHEDRAL SYMMETRY 29
  30. 30. 30
  31. 31. 31
  32. 32. 32 Adenovirus
  33. 33. 33
  34. 34. 34
  35. 35. 35 Adenovirus
  36. 36. 36
  37. 37. ICOSAHEDRAL SYMMETRY 37
  38. 38. SYMMETRY OF NUCLEOCAPSID  ICOSAHEDRAL  HELICAL 38
  39. 39. TOBACCO MOSAIC VIRUS 39 adapted from: Klug and Caspar Adv. Virus Res. 7:225
  40. 40. 40
  41. 41. Helical symmetry  Length controlled by nucleic acid  Helix may be stiff or flexible 41
  42. 42. 42
  43. 43. 43
  44. 44. COMPLEX SYMMETRY 44 POXVIRUS FAMILY
  45. 45. ENVELOPE  OBTAINED BY BUDDING THROUGH A CELLULAR MEMBRANE (except poxviruses)  POSSIBILITY OF EXITING CELL WITHOUT KILLING IT  CONTAINS AT LEAST ONE VIRALLY CODED PROTEIN 45
  46. 46. 46
  47. 47. ENVELOPE  OBTAINED BY BUDDING THROUGH A CELLULAR MEMBRANE (except poxviruses)  POSSIBILITY OF EXITING CELL WITHOUT KILLING IT  CONTAINS AT LEAST ONE VIRALLY CODED PROTEIN  ATTACHMENT PROTEIN  LOSS OF ENVELOPE RESULTS IN LOSS OF INFECTIVITY 47
  48. 48. ENVELOPE 48
  49. 49. 5 BASIC TYPES OF VIRAL STRUCTURE 49 HELICAL ENVELOPED HELICAL ENVELOPED ICOSAHEDRAL COMPLEX ICOSAHEDRAL Adapted from Schaechter et al., Mechanisms of Microbial Disease nucleocapsidicosahedral nucleocapsid nucleocapsid helical nucleocapsid lipid bilayer lipid bilayer glycoprotein spikes = peplomers
  50. 50. UNCONVENTIONAL AGENTS  VIROIDS  RNA only  Small genome  Do not code for protein  So far, only known viroids are in plants 50 • hepatitis delta agent - some viroid, some virus features
  51. 51. UNCONVENTIONAL AGENTS  PRIONS  protein only?  do not contain any nucleic acid? 51
  52. 52. LIVING OR DEAD? 52
  53. 53. CLASSIFICATION  BASIC STRUCTURE AND MOLECULAR BIOLOGY  particularly important as diagnostic and therapeutic abilities expand  past schemes  host range  tissue infected  type of cell infected  mode of transmission  disease caused 53 Arboviruses (arthropod borne)
  54. 54. CLASSIFICATION  NUCLEIC ACID  CAPSID  PRESENCE OF ENVELOPE  REPLICATION STRATEGY 54
  55. 55. CLASSIFICATION NUCLEIC ACID  RNA or DNA  segmented or non-segmented  linear or circular  single-stranded or double-stranded  if single-stranded  is genome mRNA (+) sense or complementary to mRNA (-) sense 55
  56. 56.  symmetry  icosahedral, helical, complex  enveloped or non-enveloped  number of capsomers 56 CLASSIFICATION CAPSID
  57. 57. CLASSIFICATION  ENVELOPE  REPLICATION STRATEGY 57
  58. 58. 58 HERPESVIRIDAE HEPADNAVIRIDAE ENVELOPED PAPILLOMAVIRIDAE POLYOMAVIRIDAE (formerly grouped together as the PAPOVAVIRIDAE) CIRCULAR ADENOVIRIDAE LINEAR NON-ENVELOPED DOUBLE STRANDED PARVOVIRIDAE SINGLE STRANDED NON-ENVELOPED POXVIRIDAE COMPLEX ENVELOPED DNA VIRUSES Modified from Volk et al., Essentials of Medical Microbiology, 4th Ed. 1991 All families shown are icosahedral except for poxviruses
  59. 59. 59 FLAVIVIRIDAE TOGAVIRIDAE RETROVIRIDAE ICOSAHEDRAL CORONAVIRIDAE HELICAL ENVELOPED ICOSAHEDRAL PICORNAVIRIDAE CALICIVIRIDAE NONENVELOPED SINGLE STRANDED positive sense BUNYAVIRIDAE ARENAVIRIDAE ORTHOMYXOVIRIDAE PARAMYXOVIRIDAE RHABDOVIRIDAE FILOVIRIDAE SINGLE STRANDED negative sense REOVIRIDAE DOUBLE STRANDED RNA VIRUSES ENVELOPED HELICAL ICOSAHEDRAL NONENVELOPED Modified from Volk et al., Essentials of Medical Microbiology, 4th Ed. 1991
  60. 60. Adenovirus
  61. 61. Herpesviridae
  62. 62. Influenza Virus
  63. 63. Smallpox Virus
  64. 64. Virus Classification  Historically based on:  Host preference: Plant, insect, animal, human  Target organ: respiratory, hepatic, enteric, etc.  Vector: arboviruses  Overlapping, inconsistent  Currently based on molecular biology of genome and biophysical structure
  65. 65. Virus Classification  Viruses with similar structural, genomic & replication properties are grouped into families (suffix: viridae) e.g. Herpesviridae  Families subdivided into genera (suffix: virus) e.g. Herpes simplex virus, Cytomegalovirus, Varicella zoster virus  Subtypes based on nucleotide sequence and antigenic reactivities e.g. Herpes simplex virus type 1, Herpes simplex virus type 2
  66. 66. Virus Classification Viruses Nucleic acid: DNA RNA Envelope: Yes No Symmetry: Cubic Helical (Icosahedral) (Cylindrical)
  67. 67. Classification of Some Common Viruses Family Viruses Type of Nucleic Acid Envelope Capsid Symmetry Picornaviridae Enteroviruses, polio, hep. A ss (+) RNA No I Caliciviridae Norwalk virus ss (+) RNA No I Togaviridae Rubella ss (+) RNA Yes I Rhabodoviridae Rabies ss (+) RNA Yes H Paramyxoviridae Parainfluenza, RSV, measles, mumps ss (-) RNA Yes H Orthomyxoviridae Influenza ss (-) RNA Yes H Retroviridae HIV 1,2, HTL I,II ss (+) RNA Yes I Hepadnaviridae Hepatitis B ds DNA Yes Unknown Parvoviridae Parovirus B - 19 ss (+) or (-) DNA No I Adenoviridae Adenovirus ds DNA No I Herpesviridae HSV, CMV, EBV, VZV, HHV6 ds DNA Yes I I = icosahedral, H = helical
  68. 68. Virus Classification (Common) DNA RNA Hepatitis B Human Papilloma Virus Parvovirus B19 Adenovirus Herpesviridae Polyomaviruses Influenza RSV Parainfluenza Hepatitis A, C, D, E Enteroviruses Encephalitis viruses Measles, Mumps, Rubella Norwalk, Rotavirus Virtually all others
  69. 69. BASIC STEPS IN VIRAL LIFE CYCLE  ADSORPTION  PENETRATION  UNCOATING AND ECLIPSE  SYNTHESIS OF VIRAL NUCLEIC ACID AND PROTEIN  ASSEMBLY (maturation)  RELEASE 69
  70. 70. ADSORPTION 70
  71. 71. ADSORPTION  TEMPERATURE INDEPENDENT  REQUIRES VIRAL ATTACHMENT PROTEIN  CELLULAR RECEPTORS 71
  72. 72. - ENVELOPED VIRUSES •FUSION WITH PLASMA MEMBRANE •ENTRY VIA ENDOSOMES 72
  73. 73. PENETRATION 73herpesviruses, paramyxoviruses, HIV
  74. 74. - ENVELOPED VIRUSES •FUSION WITH PLASMA MEMBRANE •ENTRY VIA ENDOSOMES, FUSION WITH ACIDIC ENDOSOME MEMBRANE 74
  75. 75. 75
  76. 76. 76
  77. 77. PENETRATION - ENVELOPED VIRUSES 77 from Schaechter et al, Mechanisms of Microbial Disease, 3rd ed, 1998
  78. 78. VIRUS UPTAKE VIA ENDOSOMES  CALLED  VIROPEXIS / ENDOCYTOSIS / PINOCYTOSIS 78
  79. 79. PENETRATION NON-ENVELOPED VIRUSES 79
  80. 80. PENETRATION NON-ENVELOPED VIRUSES 80 entry directly across plasma membrane:
  81. 81. 81
  82. 82. UNCOATING  NEED TO MAKE GENOME AVAILABLE  ONCE UNCOATING OCCURS, ENTER ECLIPSE PHASE  ECLIPSE PHASE LASTS UNTIL FIRST NEW VIRUS PARTICLE FORMED 82
  83. 83. SYNTHESIS OF VIRAL NUCLEIC ACID AND PROTEIN  MANY STRATEGIES  NUCLEIC ACID MAY BE MADE IN NUCLEUS OR CYTOPLASM  PROTEIN SYNTHESIS IS ALWAYS IN THE CYTOPLASM 83
  84. 84. ASSEMBLY AND MATURATION  NUCLEUS  CYTOPLASM  AT MEMBRANE 84
  85. 85. smallpox virus cytoplasmic assembly and maturation 85 F. A. Murphy, School of Veterinary Medicine, University of California, Davis. http://www.vetnet.ucdavis.edu/fam_graphics/download.html
  86. 86. RELEASE  LYSIS  BUDDING THROUGH PLASMA MEMBRANE  NOT EVERY RELEASED VIRION IS INFECTIOUS 86
  87. 87. HIV budding and maturation 87Hsiung, GD et al., Diagnostic Virology 1994 p204 (D. Medina)
  88. 88. HIV – mature form 88 Briggs JA et al. Structure. (2006) 14:15-20
  89. 89. Viral Replication i) adsorption (attachment) ii) entry iii) uncoating iv) transcription v) synthesis of virus components vi) assembly vii) release
  90. 90. Viral Replication i) Adsorption (attachment):  random collision  interaction between specific proteins on viral surface and specific receptors on target cell membrane (tropism)  not all cells carrying a receptor for a particular virus can be productively infected by that virus
  91. 91. Viral Replication i) Adsorption (attachment): some viruses may use more than one host cell receptor (e.g. HIV) able to infect a limited spectrum of cell types (host range) most neutralizing antibodies are specific for virion attachment proteins
  92. 92. Viral Replication ii) Entry (penetration): 2 mechanisms - endocytosis - fusion of virus envelope with cell membrane iii) Uncoating: release of viral genome cell enzymes (lysosomes) strip off the virus protein coat virion can no longer be detected; known as the “eclipse period”
  93. 93. Viral Replication iv) Transcription/Translation/Synthesis: a) DNA viruses: • replicate their DNA in host cell nucleus mediated by viral enzymes • synthesize capsid and other proteins in cytoplasm using host cell enzymes • new viral proteins move to nucleus where they combine with new DNA to form new viruses • Exception - Poxviruses synthesize their parts in host cell’s cytoplasm
  94. 94. Viral Replication iv) Transcription/Translation/Synthesi s: b) RNA viruses: –“+” sense RNA acts as mRNA - viral proteins are made immediately in cytoplasm mediated by viral enzymes –“-” sense RNA (e.g. influenza) - lst makes a “+” sense RNA copy via viral enzyme
  95. 95. Viral Replication iv) Transcription/Translation/Synthesis: • Retroviridae (e.g. HIV) • Contain enzyme “Reverse transcriptase” • “+” sense Viral RNA  cDNA  integrated into host cell chromosome • mRNA (for viral proteins) and progeny virion RNA are synthesized from integrated viral DNA by host cell enzymes (RNA polymerases)
  96. 96. Viral Replication v) Synthesis: Protein synthesis - 2 types • structural • non-structural (enzymes for replication) Nucleic acid synthesis • new virus genome • most often by a virus - coded polymerase or replicase; with some DNA viruses a cell enzyme carries this out
  97. 97. Viral Replication vi) Assembly: may take place in cell nucleus, cytoplasm or (with most enveloped viruses) at the plasma membrane vii) Release: sudden rupture of cell gradual extrusion (budding) of enveloped viruses through the cell membrane may occur together with assembly
  98. 98. Enveloped Virus Entry via Fusion
  99. 99. Non-enveloped Virus Entry via Endocytosis
  100. 100. Outcome of Viral Infections Virus-host cell interaction may result in: 1. Cell death (lytic) - due to cytopathic effect of virus 2. Cell transformation - cell converted to malignant or cancerous cell 3. Latent infection (occult) - persistent infection in quiescent state which may reactive anytime to produce disease; continuous or intermittent shedding 4. Cell fusion to form multinucleated cells
  101. 101. Persistent Viral Infections 3 types of persistent viral infection (some overlap): 1. Chronic carrier - eg. Hepatitis B; results in chronic illness 2. Latent infection - eg. Herpesviridae; result in symptomatic or asymptomatic shedding 3. Slow virus infections - due to prolonged incubation period (eg. Measles virus and SSPE)
  102. 102. Host - Organism Relationship • Interaction between host and organism affecting the development and outcome of an infection includes: – Host’s primary physical barriers – Host’s immunologic ability to control and eliminate the invading organisms – Organism’s ability to evade, destruction/virulence – Ability of organism to spread in the body
  103. 103. Virulence of Viruses & Evasion of the Immune Response  Poorly understood processes:  Antigenic variation  Some viruses encode receptors for various mediators of immunity (eg. IL1 & TNF) thus blocking their ability to interact with receptors on their intended targets  Some viruses (eg. HIV) reduce expression of class I MHC proteins, thus reducing ability of cytotoxic T cells to kill the virus-infected cells  Direct cell-to-cell propagation  Attenuated viruses (eg. Vaccine strains)
  104. 104. Definitions  Exposure: contact with a potentially infectious agent  Infection: persistence on or within another living organism  Disease: end product (damage) resulting from an infectious process  Incubation: time from infection to development of symptoms / disease
  105. 105. Virus: Incubation Times Hours to 1-2 days:  Respiratory viruses  GI viruses 1 to 3 weeks:  Measles/Mumps/Rubel la  VZV, HSV  Chlamydia  Enteroviruses, Polio  WNV Weeks to months: • Hepatitis viruses • HIV • EBV • Rabies Months to years: • Prions
  106. 106. Routes of Transmission  Horizontal transmission:  Direct contact (secretions, blood etc.)  Respiratory (aerosol)  Contaminated inanimate objects  Insect vector (mosquitoes, ticks, etc.)  Zoonoses  Vertical transmission:  Mother to fetus [Transplacental (Congenital), Perinatally]
  107. 107. Viruses - Transmission  Can occur - with or without disease - during asymptomatic shedding - during incubation period  Transmission results in primary infection  disease; reactivation results in secondary disease
  108. 108. Viruses - Epidemiology  mode of transmission  age  gender  ethnic background / country of origin  travel history  occupation  season  underlying medical condition(s)
  109. 109. DEFINITIONS - VIRAL PROTEINS  STRUCTURAL PROTEINS  ALL PROTEINS IN A MATURE VIRION  NON-STRUCTURAL PROTEINS  VIRALLY CODED PROTEINS WHICH ARE NOT PACKAGED IN THE VIRION 111
  110. 110. EFFECTS ON HOST  MAY INHIBIT HOST DNA, RNA OR PROTEIN SYNTHESIS  DETAILS AND MECHANISM VARY 112
  111. 111. CYTOPATHIC EFFECT  ANY DETECTABLE CHANGES IN THE HOST CELL  MORPHOLOGICAL CHANGES 113
  112. 112. 114 Hockley et al. J Gen Virol 69:2455-2469 uninfected HIV infected HIV infected (at higher magnifcation)
  113. 113. CYTOPATHIC EFFECT  ANY DETECTABLE CHANGES IN THE HOST CELL  MORPHOLOGICAL CHANGES  DEATH  APOPTOSIS  INDEFINITE GROWTH 115
  114. 114. 116
  115. 115. 117
  116. 116. tissue culture cells 118epithelial epithelioid fibroblastic slides from CDC
  117. 117. epithelial cells - adenovirus 119uninfected early infection late infection slides from CDC
  118. 118. epithelial cells - respiratory syncytial virus 120uninfected respiratory syncytial virus slides from CDC
  119. 119. fibroblastic cells - herpes simplex virus 121uninfected early infection late infection slides from CDC
  120. 120. fibroblastic cells - poliovirus 122uninfected early infection late infection slides from CDC
  121. 121. 123 PLAQUE ASSAYPLAQUE ASSAY
  122. 122. 124 PLAQUE ASSAYPLAQUE ASSAY
  123. 123. 125 PLAQUE ASSAYPLAQUE ASSAY
  124. 124. 126Diluted 10 fold Diluted 100 fold Diluted 1000 fold
  125. 125. PLAQUE FORMING UNIT P.F.U. pfu 127
  126. 126. SOME POINTS TO REMEMBER  INFECTIVITY  NOT EVERY RELEASED PARTICLE IS INFECTIOUS 128 • ASSAYS – detect every particle (e.g. electron microscope) – detect infectious particles only (e.g. plaque assay)
  127. 127. GLOSSARY 129

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